Micrograph of Bradyrhizobium japonicum. B. japonicum is a gram negative, rod-shaped, nitrogen-fixing bacterium which develops
a symbiosis with the soybean plant Glycine max. B. japonicum belongs to the family, Rhizobiaceae which includes other nitrogen-fixing
bacteria that develop symbiosis with legumes. This photo shows an
individual cell and groups of cells in a characteristic aster formation.

What do
the individual partners in this symbiosis get from each other? The
plant provides the bacterium with a "safe" environment and a steady
supply of carbon for energy and growth. This carbon source is referred
to as photosynthate as it is derived from the product of photosynthesis.
In most rhizobium/legume symbiosis, photosynthate refers to the dicarboxylic
acids, succinate, fumarate and malate. In return, the bacteria provide
the plant with fixed nitrogen - nitrogen gas which has been reduced
and converted into a form readily utilized by the plant. The result
of this symbiosis is a dramatic increase in plant production without
the need for adding external fertilizer.

Source:
M.L. Guerinot, Dartmouth College

The bacterium
"communicates" with the host plant and begins a process of plant and
bacterial development that leads to a symbiotic partnership. The bacterium
will attach to root hairs and release compounds that cause the roothairs
to curl. Coordination of bacterial multiplication and inward directed
growth of the root hair results in formation of an infection thread
(a tube derived from plant membranes). It is through this infection
thread that the bacteria enter the cortical cells of the root and
begin to colonize the developing root nodule.

Source:
F. Dazzo, Michigan State University

Within
the developing root nodule, bacteria divide and begin to differentiate
into a bacteroid (a term used to refer to the bacterium existing in
a symbiotic relationship to distinguish it from the free-living bacterium),
that is capable of fixing nitrogen. The bacteroids are located inside
a structure refered to as a symbiosome that is derived from plant
membrane. One to several bacteroids can be found in a single symbiosome.
Therefore, nutrients must traverse multiple membranes to reach the
bacteroids and fixed nitrogen must follow a similar complex path to
reach the plant tissue.

Source:
M.L. Guerinot, Dartmouth College

The
nodule which results from this process is a highly specialized structure.
It provides a physical barrier which keeps the free oxygen concentration
low.

Source:
Unknown

The plant
cells within the nodule produce leghemoglobin which serves as an oxygen
carrier to the bacteria within the nodule. This enables the bacteria
to obtain enough oxygen for respiration but ensures that the oxygen
is in a bound form so that it cannot harm nitrogen fixing enzymes
inside the bacteria. Cutting open a nodule reveals the deep red color
typical of leghemoglobin when it binds oxygen.

Source:
M.L. Guerinot, Dartmouth College

Soybeans
are an important crop throughout the world and strategies to
improve crop production by manipulating the nitrogen fixing
partner are desirable.

Nitrogen fixation requires
a large input of energy and reducing potential. As a strictly respiratory
organism B. japonicum must get that energy from the respiratory
chain by oxidizing energy rich substrates and reducing oxygen. One
strategy to improve crop production is to increase the nitrogen
fixing capacity of B. japonicum through manipulation of the
respiratory chain.

B. japonicum presents
an excellent model organism for studying respiratory enzymes. There
is a large body of information describing the respiratory chains
in B. japonicum. However, other than the terminal oxidases,
little information is available on the structure, function and regulation
of the enzymes of the B. japonicum respiratory chain. B.
japonicum can express a number of terminal oxidases. Specific
terminal oxidases are expressed depending upon oxygen availability
and/or association with the symbiotic partner. In association with
soybean B. japonicum undergoes conversion to a pleomorphic
form referred to as a bacteroid. Changes in the respiratory chain
of the bacteroid are reflected in the amounts and types of cytochromes
present. In addition to the expression of alternative terminal oxidases,
the bacteroids must also express a number of other cytochromes necessary
for utilization of the nutrients available insside the nodule. The
dicarboxylic acids succinate and malate serve as energy rich substrates
for the bacteroids. These compounds can be used to provide reducing
potential to the respiratory chain of the bacterium and the enzymes
involved in conserving this reducing potential contain cytochromes..

Part of the work in our
lab focuses on the enzymes involved in transfering electrons from
the dicarboxylic acids to the respiratory
chain. We are looking at two respiratory complexes, NADH dehydrogenase
(NADH ubiquinone oxidoreductase - Nuo) and succinate dehydrogenase
(Sdh) (referred to as complexes I and II, respectively, in mitochondria).
NADH dehydrogenase takes electrons from NADH and transfers them
to ubiquinone, a lipid soluble electron carrier. Possible major
sources of NADH in bacteroids are one of the malic enzymes and malate
dehydrogenase. Both of these ezymes get electrons from malate one
of the most abundant dicarboxylic acids in the soybean nodule. Succinate
is another dicarboxylic acid that is very abundant in nodules and
the enzyme succinate dehydrogenase takes electrons from succinate
and transfers them to ubiquinone.

An additional project
in our lab is directed at determining factors that may influence
the ability of inoculant strains to compete with native strains
for infection of the roothair. One factor would be acyl homoserine
lactone (AHL) molecules that are involved in density dependent gene
expression - also known as "quorum
sensing". Another factor would be inhibitors that would
block the growth of native strains but not inoculants. To accomplish
this, we are using an inhibitor of succinate dehydrogenase (carboxin
- Vitavax) and creating carboxin resistant inoculant strains.

Yet another project in
our lab is investigating factors that influence expression of the
respiratory enzymes. Factors that we are examining are iron, heme,
oxygen and the dicarboxylic acids. Projects include studies of the
expression of the hemA, sdhCDAB and nuo genes.

Check out my research
interests to find out more about my current research involving B. japonicum.